During neural development, growing axons come to sites where they must negotiate changes in course. Using the developing mouse visual system as a model, the proposed research will examine the growth and directionality of retinal axons in the optic chiasm, a classic example of a site where axons redistribute, some remaining on the same side of the brain and others crossing. This plan results in the formation topographic maps that subserve binocular vision. First, the detailed morphology of retinal ganglion cell axon growth cones will be examined in fixed preparations within the retinofugal pathway after labeling with the labels HRP or Di-I. Emphasis will be placed on defining the trajectory and growth cone shape of ipsi- and contralaterally projecting axons within the optic chiasm. Identified dye-labeled axons will be studied ultrastructurally to investigate the neuronal and non- neuronal cellular relationships within the chiasm. Second, dye-labeled axons will be viewed in semi-intact tissue slices with conventional fluorescent optics and SIT camera or with a confocal microscope and image analysis. This will allow analysis of the dynamics of growth cone behavior within the optic chiasm in real time. Third, an in vitro assay system will be developed by which to test factors responsible for fiber rearrangement and changes in directionality within the optic chiasm. Such an assay will test whether guidance cues are associated with intrinsic cells of the optic chiasm, and whether they are membrane-bound. Retinal explants will be cocultured with cells from the chiasm, or grown on membranes derived from parts of the chiasm. The behavior of growth cones, including advance, retraction and turning behavior, will be documented with video time-lapse microscopy. These experiments aim to define the factors responsible for guidance of axons towards targets during the formation of specific topographic connections. The in vitro assay will be used in the future to test for molecular mechanisms of growth cone guidance.